Abstract

E. coli DNA gyrase uses the energy of ATP hydrolysis to introduce essential negative supercoils into the genome, thereby working against the mechanical stresses that accumulate in supercoiled DNA. Using a magnetic-tweezers assay, we demonstrate that small changes in force and torque can switch gyrase among three distinct modes of activity. Under low mechanical stress, gyrase introduces negative supercoils by a mechanism that depends on DNA wrapping. Elevated tension or positive torque suppresses DNA wrapping, revealing a second mode of activity that resembles the activity of topoisomerase IV. This 'distal T-segment capture' mode results in active relaxation of left-handed braids and positive supercoils. A third mode is responsible for the ATP-independent relaxation of negative supercoils. We present a branched kinetic model that quantitatively accounts for all of our single-molecule results and agrees with existing biochemical data.

1. Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.
2. Deparment of Physics, University of California, Berkeley, California 94720, USA.
3. John Innes Centre, Norwich NR4 7UH, UK.
4. Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.
5. Howard Hughes Medical Institute, USA.
6. Present addresses: Department of Bioengineering, Stanford University, Stanford, California 94305, USA (Z.B.), Department of Physics, Korea University, Seoul, South Korea (S.C.H.), Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138, USA (M.D.S.), and Department of Physics, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA (J.G.).
7. Deceased.
8. These authors contributed equally to this work.

Correspondence to: Carlos Bustamante^1,2,4,5 e-mail: carlos@alice.berkeley.edu

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